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Abstract
The aim of this study is to produce transgenic ovine spermatozoa carrying either bovine or ovine growth hormone (GH) cDNA sequence of Egyptian x Holstein cows or Ossimi sheep. This study has been conducted as an introduction to produce either bovine or ovine GH-transgenic sheep using sperm mediated gene transfer (SMGT) technique to improve their production. The mRNA of pituitary gland has been isolated and reversed to cDNA which cloned into pTZ57R/T cloning vector and then into pmKate2-N expression vector. The GH gene of Egyptian x Holstein crossbred (EH_GH) and Ossimi sheep breed (Os_GH) has been sequenced and characterized in this study. A total of 160 ol ovine sperm cell suspensions (40 x 10⁶ final concentrations) have been incubated with 200 ng of pmKate2-N expression vector. Total of nine experiments have been conducted; four groups for Os_GH-pmkate2-N vector, four groups for EH_GH-pmkate2-N vector and control as following; 1). Control, ovine spermatozoa have been incubated in the absence of pmKate2 vector at 37{u00B0}C for 1h., 2). 37{u00B0}C for 1h group, ovine spermatozoa have been incubated with pmkate2-N vector at 37{u00B0}C for 1h. 3). Heat shock group, ovine spermatozoa have been incubated with pmkate2-N vector at 4{u00B0}C for 20 min and then 42{u00B0}C for 2 min. 4). Heat shock + DMSO group, ovine spermatozoa have been supplemented with 3% DMSO and incubated with pmkate2-N vector at 4{u00B0}C for 20 min and then 42{u25E6}C for 2 min. 5). DMSO group, ovine spermatozoa have been supplemented with 3% DMSO and incubated with pmkate2-N vector at 37{u00B0}C for 1h. The complete coding sequences (CDS) of EH_GH and Os_GH were detected, annotated comprehensively and registered in Gene Bank under accession number AC: KP221576 and Ac: KP221575, respectively